X-linked hypophosphatemia (XLH), the most common inherited form of rickets in North America, is an X-linked dominant Mendelian disorder characterized by hypophosphatemia from renal Pi wasting, abnormal vitamin D metabolism, and impaired mineralization of bone. As a disease model to illuminate our understanding on bone and kidney functions, XLH has been studied extensively particularly after its experimental animal, i.e., the hypophosphatemic (Hyp) mouse, became available. In recent years, significant advances have been made on this disorder, which include the discovery of PHEX/Phex gene mutations in XLH and the realization that primary defects in bone may lead to the release of factor(s) causing bone mineralization defect and renal Pi wasting. However, the identity of these bone- derived pathogenic factors and the exact role of PHEX/Phex mutations in the release of these factors remain unknown. Aiming to identify these bone derived pathogenic factors, we have identified two Hyp-specific proteins from the conditioned culture media derived from in vitro cultured Hyp mouse osteoblast cells (OC). One is the cathepsin D (Cat D) capable of inhibiting OC mineralization, and the other one is the 14-3-3-gamma protein capable of inhibiting proximal tubular cell Pi uptake. Our preliminary studies using respective inhibitors further supported their pathogenic roles in Hyp mice. Since we have also found in Hyp mouse sera a higher level of 14-3-3-gamma protein but not Cat D, we hypothesize that the increased release of Cat D by Hyp mouse OC constitutes a local autocrine/paracrine system in bones contributing to the Hyp mouse bone defect, whereas the increased release of 14-3-3-gamma protein by Hyp mouse OC may serve as a circulating phosphaturic factor contributing to the Hyp mouse kidney defect. The aims of our present proposal are two fold: (1) to further characterize the pathogenic roles of Cat D and 14- 3-3-gamma protein in Hyp mice, and (2) to examine the role of Phex defect in the release of these osteoblast-derived pathogenic factors in Hyp mice.